Abstract
Purpose :
Macula-specific enrichment of carotenoids is thought to be a result of the action of carotenoid transport proteins and carotenoid binding proteins. CD36 is a cell surface glycoprotein capable of many different functions in the body. It is a class B scavenger receptor that mediates HDL uptake. Deficiency of this protein results in defective fatty acid and oxidized lipid uptake. It is known that lutein and zeaxanthin are transported in serum on lipoproteins. Cell culture studies using ARPE 19 cells expressing SRB1, another scavenger receptor protein, have shown that knock down of this gene can result in a decrease in cellular uptake of carotenoids. CD36, rather than SRB1, is robustly expressed in primate retina. In addition, studies on silkworm have suggested that a CD36 homolog is involved in the transport of lutein. Using the results from these studies, we hypothesize that CD36 may function to transport carotenoids into the retina.
Methods :
Monkey (Macacca mulatta) eyes from 4-6 year old animals and mice eyes from 6 month old C57BL/6 animals were enucleated, fixed in 4% paraformaldehyde overnight, and cryoprotected in sucrose gradient prior to embedding in OCT. 10-12um sections were obtained by cryosectioning. Sections were stained with antibodies against CD36 (1:500) and/or RPE65 (1:1000), and glutamine synthetase (1:5000). Confocal images were obtained at 40X magnification. Retina, RPE, and macula tissues were isolated from human donors (ages 54, 72). Total RNA and total protein were isolated. RT-PCR and Western Blot were conducted to determine the presence of CD36 in these tissues. Using recombinant CD36 protein, we conducted surface plasmon resonance (SPR) studies to determine whether this protein can bind macular carotenoids.
Results :
Our data show that CD36 transcripts and protein are present in the RPE and retinal layers in mice, monkeys, and humans. Macular sections of monkey eyes show strong co-localization of CD36 with RPE65, an RPE marker. SPR studies show that CD36 can bind lutein and meso-zeaxanthin with moderate affinity, characteristic of a transport protein.
Conclusions :
Our data point to the significance of CD36 in carotenoid transport across vertebrate species. Since lutein and zeaxanthin are enriched in the macular region of humans, identifying the role of CD36 mediated transport of these carotenoids may further enhance our understanding of processes that may go awry in age-related macular degeneration.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.